1. Field of the Invention
The invention relates generally to molecularly-imprinted material and in particular to molecularly-imprinted material made by template-directed synthesis.
2. Description of the Related Art
Enzymes are commonly exploited for practical uses, including as catalysts in synthetic processes, as detection reagents in chemical and biological sensors, and as catalysts in decontamination of environmental pollutants and other toxic agents. Their usefulness is largely due to their exquisite functional selectivity and regio- and stereospecificity. However, the usefulness of enzymes for practical purposes is limited by their intolerance to harsh conditions, particularly to conditions involving nonaqueous environments, temperature extremes, or the presence of materials that are toxic to the enzyme. In addition, enzymes may have a short shelf-life under ambient conditions and may require refrigerated storage to remain active.
Similarly, antibodies are useful for practical purposes such as for detecting or separating specific materials in complex mixtures. As with enzymes, the usefulness of antibodies is due to their functional selectivity and regio- and stereospecificity. Also, as with enzymes, their usefulness is limited by their intolerance for harsh conditions.
As a result of the difficulties in exploiting enzymes and antibodies on a large scale and in harsh environments, efforts have been made to develop enzyme and antibody mimics, that is, materials that can function as enzymes or antibodies, but which have a more durable composition. Specifically, efforts have been made to utilize the principles of molecular recognition to create artificial enzyme active sites or antibody binding sites through molecular imprinting of enzyme transition state analogs or antibody antigens in polymers and inorganic matrices. For example, molecularly printed materials are described in the following patents and publications incorporated herein by reference: U.S. Pat. No. 5,110,833 to Mosbach; U.S. Pat. No. 5,310,648 to Arnold et al; U.S. Pat. No. 5,372,719 to Afeyan et al; U.S. Pat. No. 5,453,199 to Afeyan et al; U.S. Pat. No. 5,461,175 to Fischer et al; U.S. Pat. No. 5,587,273 to Yan et al; U.S. Pat. No. 5,630,978 to Domb; U.S. Pat. No. 5,641,539 to Afeyan et al; U.S. Pat. No. 5,728,296 to Hjertxc3xa9n et al; U.S. Pat. No. 5,750,065 to Kilbane II; U.S. Pat. No. 5,756,717 to Paliwal et al; U.S. Pat. No. 5,786,428 to Arnold et al; U.S. Pat. No. 5,814,223 to Hjertxc3xa9n et al; U.S. Pat. No. 5,821,311 to Mosbach et al; U.S. Pat. No. 5,858,296 to Domb; U.S. Pat. No. 5,872,198 to Mosbach et al.; Mosbach, K. et al, xe2x80x9cThe Emerging Technique of Molecular imprinting and Its Future Impact on Biotechnologyxe2x80x9d, Biotechnology, vol 14, February 1996, pp 163-170; G. Wulff, xe2x80x9cMolecular Imprinting in Cross-Linked Materials with the Aid of Molecular Templatesxe2x80x94A Way towards Artificial Antibodiesxe2x80x9d Angew. Chem. Intl. Ed. Engl., 34, 1812-1832 (1995); P. Hollinger, et al., xe2x80x9cMimicking Nature and Beyondxe2x80x9d Trends in Biochemistry, 13(1), 7 9 (1995); Haupt, K., Mosbach, K. Trends Biotech, 16, 468-475 (1997); Davis et al, xe2x80x9cRational Catalyst Design via Imprinted Nanostructured Materialsxe2x80x9d Chem. Mater. 8 (1996) pp 1820-1839. and Wulff. G. et al, xe2x80x9cEnzyme models Based on Molecularly Imprinted Polymers with Strong Esterase Activityxe2x80x9d Angew. Chem. Int. Ed. Engl., 36 1962 (1997).
During a typical imprinting process, a molecule to be imprinted is combined with a mixture of functionalized and non-functionalized monomers so that the monomers surround the molecule to be imprinted. In the process, functionalized monomers align themselves in a binding relationship to complementary functional groups on the imprint molecule. The monomers are then polymerized, thereby encasing the imprint molecule within the polymer. The imprint molecule is then washed away, and the resulting material contains imprinted binding sites which are the xe2x80x9cnegativexe2x80x9d of the imprint molecule. The complementary binding groups, arising from the functionalized polymer groups incorporated during the imprinting, are specifically positioned to enhance the preferential substrate binding and, if desired, subsequent catalysis.
To date, the methods of molecular imprinting described above have achieved only modest success in producing imprinted materials that exhibit selectivity and catalytic activity. The reason for this is that in order to be effective in wide scale use, antibody and enzyme mimics must have binding/active sites that are nearly homogeneous (in specificity and activity), well formed (based on shape and reactivity), and easily accessed by reactant molecules (access is affected by shape, size and polarity of the channels leading to the catalytic site). Site homogeneity and site accessibility are both equally important. The imprinted sites created by currently known methods are generally not very accessible and are generally not homogeneous, that is, they often have different binding affinities and/or reactivities. These problems arise from the methods used in forming the polymer imprint and providing access to the binding sites. Using the conventional imprinting process, the imprinted sites are completely encased within the polymer. In order to enable access to the sites, the polymer may be ground up, thereby exposing the sites. However, doing so causes the deformation of a large number of the binding sites and irreversibly alters the shape-specificity and the complementary binding of the sites, thereby adversely affecting their selectivity and activity. In an alternative method of enabling access to the imprinted sites, porogens (typically inert solvents) may be incorporated among the polymerizable monomers in the imprinting process. After polymerization, the porogens are washed away, creating pores that allow access to the binding sites. However, as the porogens are removed, some of the structural integrity of the polymer is lost, leading to deformation of the sites. The resultant loss in specificity and activity is similar to that observed as a result of grinding up the polymer.
Recently, efforts have been made to improve accessibility by creating imprinted sites on silica or polymer surfaces. In general, this approach involves linking complementary hydrogen-bonding functionalized silanes to the imprint molecule and then creating the molecular recognition site by attaching this xe2x80x9cscaffoldingxe2x80x9d to the surface of a silica or polymer particle. After the imprint molecule is washed away, a binding site with affinity for specific molecules remains on the surface of the particle. This approach is described in the following publications incorporated herein by reference: Lele B. S, et al xe2x80x9cMolecularly Imprinted Polymer Mimics of Chymotrypsin 1. Cooperative Effects and Substrate Specificityxe2x80x9d React. Funct. Polym 39(1), 37-52 (1999); Lele, B. S., xe2x80x9cMolecularly Imprinted Polymer Mimics of Chymotrypsin 2. Functional Monomers and Hydrolytic Activityxe2x80x9d React. Funct. Polym 40(3), 215-229 (1999); and Hwang K-O, et al, xe2x80x9cTemplate-Assisted Assembly of Metal Binding Sites on a Silica Surfacexe2x80x9d, Mater. Sci. Eng. C, 3, 137 (1995).
This approach has some important limitations: First, the scaffolding process places the imprint molecule on the surface of the particle. Consequently, this procedure imprints only the functionality of the imprint molecule and not the molecule""s shape. Additionally, there are limits to how much of the imprint molecule""s functionality can be imprinted using this procedure. This is essentially a 2-dimensional form of imprinting in that only those functional groups of the imprint molecule with pre-attached complementary binding groups oriented towards the particle surface would be imprinted. Functional groups with pre-attached complementary binding groups oriented away from the surface would not be tethered to the surface and so would not be imprinted. The fewer functional groups imprinted, the lower the selectivity of the imprinted site for the target molecule, and the binding of the target molecule also will be much weaker. Second, because of the chemistry involved in attaching the imprint molecule-complementary groups complex to a surface, the xe2x80x9cscaffoldingxe2x80x9d procedure is limited to the imprinting of particle or planar surfaces. This procedure is not useful for imprinting porous materials due to difficulties in introducing the reactants into the pores. Even if the attachment to the surfaces of the pores could be achieved, this imprinting would necessarily restrict the flow of any target molecules through the pores, thereby creating the accessibility problems this approach was designed to alleviate.
In a separate field of technology, methods have been developed for making particles and porous materials by template-directed synthesis. In these methods, surfactants are used to create molecular microstructures such as micelles or reverse micelles in a solvent medium and then inorganic or organic monomers are polymerized around the molecular microstructures at the surfactant-solvent interface. When the surfactant is removed, the remaining material has a size and shape complementary to the size and shape of the molecular microstructures. By controlling variables such as surfactant selection and concentration, a variety of different microstructure shapes such as micellar, cubic, tetragonal, lamellar, tubular and reverse micellar can be formed and, consequently, monodisperse particles of a variety of different sizes and porous materials with a variety of different shapes of pores and channels can be created. Methods of making porous material are described, for example, in the following patents and publications incorporated herein by reference: U.S. Pat. No. 5,250,282 to Kresge et al; U.S. Pat. No. 5,304,363 to Beck et al; U.S. Pat. No. 5,321,102 to Loy et al; U.S. Pat. No. 5,538,710 to Guo et al; U.S. Pat. No. 5,622,684 to Pennavaia et al; U.S. Pat. No. 5,750,085 to Yamada et al; U.S. Pat. No. 5,795,559 to Pinnavaia et al; U.S. Pat. No. 5,786,294 to Sachtler et al; and U.S. Pat. No. 5,858,457 to Brinker et al; J. C. Vartuli, et al, xe2x80x9cEffect of Surfactant/Silica Molar ratios on the Formation of Mesoporous Molecular Sieves: Inorganic Mimicry of Surfactant Liquid-Crystal Phases and Mechanistic Implicationsxe2x80x9d Chemistry of Materials, 6, 2317 2326 (1994); C. A. Morris, et al xe2x80x9cSilica Sol as a Nanoglue: Flexible Synthesis of Composite Aerogelsxe2x80x9d Science, 284, 622-624 (1999); B. T. Holland et al, xe2x80x9cSynthesis of Highly Ordered, Three-Dimensional, Macroporous Structures of Amorphous or Crystalline Inorganic Oxides, Phosphates and Hybrid Compositesxe2x80x9d Chem Mater 11, 795-805 (1999); and M. Antonietti, et al, xe2x80x9cSynthesis of Mesoporous Silica with Large Pores and Bimodal Pore Size Distribution by Templating of Polymer Laticesxe2x80x9d Advanced Materials 10, 154-159 (1998). These materials, while being able to distinguish molecules on the basis of size, typically lack the specificity and activity of enzymes and antibodies. Methods for making monodisperse silica particles by hydrolyzing alkoxysilanes in a surfactant-stabilized water-in-oil microemulsion containing ammmonia are described, for example, in the following patents and publications incorporated herein by reference: U.S. Pat. No. 5,209,998 to Kavassalis at al; W. Stober et al, xe2x80x9cControlled Growth of Monodisperse Silica Spheres in the Micron Size Rangexe2x80x9d J. Colloid Interface Sci., 26, 62 (1968); Lindberg et al, xe2x80x9cPreparation of Silica Particles Utilizing the Sol-Gel and the Emulsion-Gel Processesxe2x80x9d Colloids and Surfaces A 99, 79 (1995); P. Espiard et al, xe2x80x9cA Novel Technique for Preparing Organophilic Silica by Water-In Oil Microemulsionsxe2x80x9d Polymer Bulletin, vol. 24, 173 (1990); H. Yamauchi et al, xe2x80x9cSurface Characterization of Ultramicro Spherical Particles of Silica Prepared by W/O Microemulsion Methodxe2x80x9d, Colloids and Surfaces, Vol 37, 71-80 (1989); Markowitz et al, xe2x80x9cSurface Acidity and Basicity of Functionalized Silica Particlesxe2x80x9d Colloids and Surfaces A: Physicochem Eng. Aspects 150, 85-94 (1999). The formation of silica gel-coated metal and semiconductor nanoclusters is described in U.S. Pat. No. 5,814,370 to Martino et al. As described below, unfunctionalized silica particles have little or no catalytic activity; catalytic activity is increased with functionalized silica particles, but not to the level achieved with imprinted materials.
Accordingly, it is an object of the present invention to provide an imprinted material and a method of making an imprinted material wherein the imprinted sites are easily accessible to target molecules.
It is a further object of the present invention to provide an imprinted material and a method of making an imprinted material wherein the imprinted sites are nearly homogeneous in activity and specificity.
These and other objects are achieved by a method for making a molecularly imprinted solid structure by the steps of: providing a surfactant compound having a headgroup, wherein the headgroup comprises a group to be molecularly imprinted; combining the surfactant compound with a solvent to form a mixture and maintaining the mixture from the previous step so that molecules of the surfactant self-assemble to form at least one supramolecular structure having exposed imprint groups; combining the mixture with at least one reactive monomer so that the supramolecular structure serves as a template for the organizing of molecules of the reactive monomer surrounding or in contact with the supramolecular structure, including surrounding the imprint groups; maintaining the mixture so that the reactive monomers react with each other to form a solid structure having a shape complementary to the shape of the supramolecular structure, including the shape of the exposed imprint groups; and removing the supramolecular structure from the solid structure. The resulting structure is a porous material or particle having molecularly imprinted sites. Because the imprinting is accomplished into the material""s surface, the imprinted sites are easily accessible to target molecules. And because no drastic processing steps such as grinding are required to achieve accessibility, the imprinted sites tend to be homogeneous in specificity and reactivity.